Rewiring Escherichia coli to transform formate into methyl groups

Biotechnological applications are steadily growing and have become an important tool to reinvent the synthesis of chemicals and pharmaceuticals for lower dependence on fossil resources. In order to sustain this progression, new feedstocks for biotechnological hosts have to be explored. One-carbon (C₁-)compounds, including formate, derived from CO₂ or organic waste are accessible in large quantities with renewable energy, making them promising candidates. Previous studies showed that introducing the formate assimilation machinery from Methylorubrum extorquens into Escherichia coli allows assimilation of formate through the C₁-tetrahydrofolate (C₁-H₄F) metabolism. Applying this route for formate assimilation, we here investigated utilisation of formate for the synthesis of value-added building blocks in E. coli using S-adenosylmethionine (SAM)-dependent methyltransferases (MT).

We first used a two-vector system to link formate assimilation and SAM-dependent methylation with three different MTs in E. coli BL21. By feeding isotopically labelled formate, methylated products with 51-81% ¹³C-labelling could be obtained without substantial changes in conversion rates. Focussing on improvement of product formation with one MT, we analysed the engineered C₁-auxotrophic E. coli strain C₁S. Screening of different formate concentrations allowed doubling of the conversion rate in comparison to the not formate-supplemented BL21 strain with a share of more than 70% formate-derived methyl groups.

Within this study transformation of formate into methyl groups is demonstrated in E. coli. Our findings support that feeding formate can improve the availability of usable C₁-compounds and, as a result, increase whole-cell methylation with engineered E. coli. Using this as a starting point, the introduction of additional auxiliary enzymes and ideas to make the system more energy-efficient are discussed for future applications.